Our project focuses on the neurobiology of Rett Syndrome (RS) to understand the pathways through which genetic insufficiency of MeCP2 protein leads to the disorder's neurobehavioral manifestations and arrest of brain growth. We have hypothesized that in RS, neurons and inter-neuronal connections are disrupted during the most dynamic phase of brain growth when synapses are being formed and eliminated. During the last period of support we focused on synaptic neurotransmitter abnormalities in human postmortem brain from girls with RS because we hypothesized that the disorder targeted the development of axono-dendritic connections. We discovered that the densities of several neurotransmitter receptors, including NMDA and AMPA-type glutamate receptors are elevated in young girls less than 10 years of age, while these receptors are depleted compared to controls in older girls. We hypothesize that these age-related biphasic changes may underlie the seizures, stereotyped hand movements and other signs of encephalopathy seen in young girls with RS. The cholinergic nucleus basalis of Meynert (nBM) is also disrupted in RS and we found that this leads to Rett-like shrinkage of neurons in developing cerebral cortex in rodents. When antibodies to MeCP2 became available, we found that expression in the brain is primarily localized to neurons, consistent with our overall hypothesis. In proposed studies, we will focus on synaptic abnormalities in "knockout" and "knockin" genetic models of MeCP2 insufficiency. Using sophisticated morphologic, molecular and cell biology approaches and behavioral testing, we will determine whether these mice exhibit phenotypic changes that resemble the pathophysiological features of RS. Our studies will focus on alterations in the cerebral cortex, nBM and/or the cerebellum, regions that show significant changes in RS. In in vitro experiments, we will explore the pathophysiology of synaptic connections and the effects of replacement of MeCP2 in cultured cerebellar and cortical neurons using transfection technology. Our studies are directly relevant to the long-term goal of this Program Project to discover ways to modify secondary neuropathological and neurobehavioral abnormalities in RS, even if it not possible to reverse the primary abnormality of transcriptional regulation.